This application addresses broad Challenge Area: (06) Enabling Technologies and specific Challenge Topic: 06-MH-103 New technologies for neuroscience research. Develop technologies for neuroscience research that are software-based, (e.g., informatics tools, implementation of data analytic algorithms), hardware-based (e.g., instrumentation or devices), or biology-based (e.g., driven by conditional gene expression or bioactive agents). Contact: Michael F. Huerta, Ph.D., 301-443-1815, mhuert1@mail.nih.gov Establishing a network diagram of the brain is one of the major challenges of modern neurobiology and medicine, particularly a diagram of genetic connectivity. Nowhere is this need more clear than for the social-emotional system, where dysregulation of circuitry has been implicated in the most devastating mental illnesses, depression, schizophrenia and autism. This proposal seeks to deploy the next generation neurobiologic technologies and novel software tools needed to generate gene-specific 3-dimensional reconstructions from serial sections of long range axon projections or wiring diagrams of the limbic system, at the axon level and on the size scale of primate brain. This software will fill two striking gaps that have seriously hindered the generation of maps of primate brain circuitry. Blocked by lack of existing software for both aligning sequential sections and for handling large datasets, current circuit reconstruction methods cannot handle axon projections that travel through more than a single, typical ~30[unreadable]m section. Therefore, although there are elegant emerging approaches to tract tracing, current technology permits only visualization of cells in a single section of a mouse hippocampus but prevents providing answers to the critical questions at the systems level. The long range links to other cortical or subcortical structures involved in regulation of emotion or cognition, particularly at the scale needed for primate systems cannot yet be visualized. Moreover, there are no programs for making the leap between axon projections and larger tracts defined by DTI or MRI. The current proposal takes advantage of a unique multidimensional collaboration among the Scientific Computing and Imaging Institute (SCI) experts in image reconstruction and three-dimensional visualization (Tasdizen, Jones) who have recently (Anderson, 2009) developed a computational framework for 3-dimensional neurocircuitry of the retina and in fluorescence image reconstruction (Roysam), experts in non-human primate brain circuitry and neuroanatomy (Angelucci, Hof), experts in animal and human DTI/ MRI (Hsu), and experts in multicolor fluorescence imaging of axon projections (Korenberg, Angelucci). This unique collaborative project will help to maintain two junior investigators and three early career investigators, and will create seven new research jobs in the fields of neurobiology and computation. The four aims will: 1) Generate high signal- to-noise multicolor fluorescence images of neuropeptide projections in the macaque limbic system (hypothalamus- a subset of limbic targets) at axon resolution, 2) Acquire and store images of more than 400 serial sections in mosaic 2 [unreadable]m stacks equal to158 terabytes, 3) Reconstruct the hypothalamic- projections found with arginine-vasopressin and a Williams syndrome gene product to a subset of limbic targets from serial images into a single continuous dataset, and 4) Automatically reconstruct axon pathways to their targets. The goal is to create and integrate this novel set of software and neurobiologic technologies that will accelerate wiring the brain of mammals. These technologies will provide the critical missing framework necessary for image acquisition, manipulation of terabyte datasets, serial section reconstruction, automatic and manual tract tracing, and gene specific wiring diagrams or connectomes at the scale of primate brains and for bridging the gap between axon projections and high resolution tracts obtained by MRI and DTI of non-human primates and humans. of this research to public health: Establishing a network diagram of the brain is one of the major challenges of modern neurobiology and medicine, particularly a diagram of genetic connectivity. Nowhere is this need clearer than for the brain system controlling social behavior and emotion, where dysregulation of circuitry has been implicated in the most devastating mental illnesses, depression, schizophrenia and autism that together affect more than 13 million Americans. Creating solutions to understand these brain systems will broadly accelerate unraveling the connectivity of many circuits that are disturbed in other neurologic diseases, including those involved in Parkinson's, Alzheimer's, and addiction. In summary, the tools we are creating will transform the field of mental illness by providing genetic links to the underlying brain circuitry. This knowledge will provide insights into new drug targets to prevent, treat, and ultimately cure mental illness.